Various analytical procedures and devices are commonly employed in assays to determine the presence and/or absence of analytes in a test sample. For instance, immunoassays utilize mechanisms of the immune systems, wherein antibodies are produced in response to the presence of antigens that are pathogenic or foreign to the organisms. These antibodies and antigens, i.e., immunoreactants, are capable of binding with one another, thereby causing a highly specific reaction mechanism that may be used to determine the presence or concentration of that particular antigen in a biological sample. There are several well-known techniques for detecting the presence of an analyte.
One such technique is described in WO 01/38873 to Zhang. Zhang describes flow-through electrochemical biosensors designed to detect the presence of an analyte. FIG. 2 of Zhang, for instance, illustrates a sensor assembly 5 that includes an absorbent pad 18, a wicking mesh 22, and a conjugate pad 20 that overlay an application area 14′ and a detection area 16′. The wicking mesh 22 functions as a carrier to deliver the fluid sample through capillary action to the detection area 16′ where the analyte will become immobilized on the electrode surface. In Example 4 of Zhang, various materials of different pore sizes (ranging from 0.63 to 100 microns) were tested to determine the time for a buffer solution to flow 4 centimeters along the membrane. The times ranged from 40 seconds to 3 minutes, 45 seconds. Zhang indicates that any of the membranes tested could be used to provide a rapid test.
Unfortunately, conventional flow-through electrochemical biosensors, such as described above, possess various problems. For instance, traditional flow-through assay devices require a large sample volume because of the presence of a large sampling pad, wicking pad, and porous membrane. Moreover, the contact of the sample with the working electrode surface is not always sufficient because a large portion of the sample does not flow through the electrode surface, rather it flows through the membrane itself and bypasses the working electrode. Furthermore, the positioning of two or more electrodes close to each other poses a challenge for the surface treatment of the working electrode. In particular, such biosensors often exhibit substantial background interference due to contamination of the counter/reference electrode(s) resulting from surface treatment of the working electrode. As such, a need still exits for an improved flow-through, electrochemical assay device.